Cellulolytic Properties of a Potentially Lignocellulose-Degrading Bacillus sp. 8E1A Strain Isolated from Bulk Soil

Jakub Dobrzyński,Barbara Wróbel,Ewa Beata Górska

doi:10.3390/agronomy12030665

Jakub Dobrzyński, Barbara Wróbel + Show 1 more

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https://doi.org/10.3390/agronomy12030665

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Abstract

Cellulolytic enzymes produced by spore-forming bacteria seem to be a potential solution to the degradation of lignocellulosic waste. In this study, several dozen bacterial spore-forming strains were isolated from soil and one of them was selected for further studies. The studied bacterial strain was identified to genus Bacillus (strain 8E1A) by 16S rRNA gene sequencing. Bacillus sp. 8E1A showed an activity of carboxymethyl cellulase (CMCase) with visualization with Congo Red-25 mm (size of clear zone). To study CMCase, filter paper hydrolase (FPase), and microcrystalline cellulose Avicel hydrolase (Avicelase) production, three different cellulose sources were used for bacterial strain cultivation: carboxymethyl cellulose (CMC), filter paper (FP), and microcrystalline cellulose Avicel. The highest CMCase (0.617 U mL−1), FPase (0.903 U mL−1), and Avicelase (0.645 U mL−1) production of Bacillus sp. 8E1A was noted for using CMC (after 216 h of incubation), Avicel cellulose (after 144 h of incubation), and CMC (after 144 h of incubation), respectively. Subsequently, the cellulases’ activity was measured at various temperatures and pH values. The optimal temperature for CMCase (0.535 U mL−1) and Avicelase (0.666 U mL−1) activity was 70 °C. However, the highest FPase (0.868 U mL−1) activity was recorded at 60 °C. The highest CMCase and Avicelase activity was recorded at pH 7.0 (0.520 and 0.507 U mL−1, respectively), and the optimum activity of FPase was noted at pH 6.0 (0.895 U mL−1). These results indicate that the cellulases produced by the Bacillus sp. 8E1A may conceivably be used for lignocellulosic waste degradation in industrial conditions.

Highlights

Every year, approximately 200 billion tons of plant biomass is produced on Earth, approximately 90% of which is lignocellulosic waste [1]
All bacterial strains were capable of decomposing D-ribose, D-galactose, D-glucose, D-fructose, D-mannitol, amygdalin, esculin, D-cellobiose, D-maltose, starch, D-melibiose, and D-sucrose
The studied isolate—Bacillus sp. 8E1A—was able to produce all studied enzymes on each of the substrates used for the culture

Summary

Introduction

Approximately 200 billion tons of plant biomass is produced on Earth, approximately 90% of which is lignocellulosic waste [1]. Lignocellulosic waste may be used for producing biofuels (e.g., bioethanol and biogas), gaining importance in the era of the depletion of conventional fossil fuel resources [2–4]. Cellulose is the most common biopolymer in nature, made of β-D-glucose molecules, linked by β-1,4-glycosidic bonds. It contains crystalline and amorphous regions [6]. Cellulolytic enzymes include: (1) endo-β-1,4-glucanases (EC 3.2.1.4), which randomly cleave β-1,4 glycosidic bonds located in the amorphous regions of the cellulose—an example of endoglucanase is carboxymethyl cellulase, (2) exo-1,4β-glucanases (EC 3.2.1.91), which detach glucose and cellobiose units from reducing or non-reducing ends of the cellulose chain, e.g., enzymes that degrade a microcrystalline cellulose Avicel-Avicelase, and (3) cellobiases (β-glucosidase), which convert cellobiose to glucose (EC 3.2.1.21) [7–9]

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